Composition in particular cosmetic or dermatological composition, containing oligosaccharides and preparation method and cosmetic treatment method

ABSTRACT

The invention concerns a composition comprising at least a first constituent which is an oligosaccharide of formula (I) containing 3 to 5 oside units and having at least a D-galactose unit bound by an α[1-6] bond with a sucrose unit. The invention is characterized in that in said sucrose unit R1, R2, R3, R4, independently of one another represent a hydrogen atom, an alkyl group containing 1 to 10 carbon atoms and optionally having at least an unsaturation, a sulphate function or else an ose and, at least a second constituent which is a molecule positively charged with a physiological pH and stimulating pinocytosis or a molecule stimulating membrane penetration.

[0001] The present invention relates to a composition, especially acosmetic or dermatological external topical composition, containingoligosaccharides, to its preparation process and to a cosmetic treatmentprocess comprising the application of a cosmetic composition.

[0002] The field of the present invention is that of unpleasant symptomspossibly associated with allergy and in particular with asthma, eczema,pruritus, psoriasis, allergic conjunctivitis, urticaria, reactions toinsect bites, and itching, in particular symptomatic itching of majorburn sufferers.

[0003] The degranulation properties of polymorphonuclear leukocytes andespecially of eosinophils, basophils and mastocytes are known to beinvolved in allergic phenomena.

[0004] Thus, mature blood basophils have granules distributed randomlyand edged with a membrane, these granules contain various products(heparin, SRS-A, ECF-A) which are released when a suitable stimulusinduces a degranulation. This stimulus is usually an allergen that pairsup with the specific IgEs bound to the surface of the cell via suitablereceptors. The products released by the degranulation are responsiblefor some of the unpleasant symptoms associated with the allergy, butthey are also involved in antiparasitic immunity.

[0005] The main compounds of the prior art known for their antiallergicproperties are compounds of the antihistamine family such aschloropheniramine (polaramine). These compounds are not without sideeffects, such as risks of drowsiness. Consequently, the presentinvention proposes the use of a combination of compounds that are freeof the side effects of standard antihistamines and that neverthelessmake it possible to treat allergies and other complaints or disorders ofthe same type.

[0006] The Applicant has demonstrated that the use of certainoligosaccharides is effective in reducing or even blocking theunpleasant symptoms possibly associated with allergy and as ananti-inflammatory.

[0007] Without wishing to be bound by any particular theory, it appearsthat the Applicant has shown that certain oligosaccharides have anantidegranulating, antigranulating or antiactivating activity on atarget cell of the allergy and especially on the human basophilstimulated according to an IgE-dependent reaction.

[0008] Other specialized skin cells (such as, for example, the neurones,which, by release of substance P, are responsible for pain but alsoinduce a degranulation of basophils) are also concerned by thisactivity.

[0009] The phenomenon of degranulation and also the unpleasant symptomsassociated therewith are known to those skilled in the art; however, theinventors of the present patent application do not rule out thepossibility that the combination used for the purposes of the presentpatent application intervenes upstream of the degranulation: during thematuration of the granules. Accordingly, according to the inventors,this combination might also have “antigranulating” activity.

[0010] The present invention more particularly relates to a combinationcomprising at least two components:

[0011] a first component that is an oligosaccharide of formula Icontaining from three to five saccharide units and having at least oneD-galactose unit linked via an α[1-6] bond with a sucrose unit:

[0012] in which R₁, R₂, R₃ and R₄, independently of each other,represent a hydrogen atom, an alkyl group containing from 1 to 10 carbonatoms and optionally having at least one unsaturation, an acid functioncontaining from 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide,

[0013] and at least one second component that is a molecule that ispositively charged at physiological pH and that promotes pinocytosisand/or a molecule that promotes membrane penetration.

[0014] The second component acts as a vector for the first component,this first component being the active principle. The essential role ofthis second component is to allow the first component, the activeprinciple, to cross the white blood cell membrane.

[0015] Said oligosaccharide may be chosen in particular from the groupconsisting of raffinose and stachyose in free or derived form. Thesecond component is preferably a saponin or a basic amino acid, and itis chosen even more preferably from the group consisting of saponins andarginine.

[0016] Preferably, the following natural saponins will be used:harpagosides, ginsenosides, saponins of quillaja saponaria QS III and QS21A type.

[0017] The present invention relates to the use of a combination of atleast two components for the manufacture of a cosmetic composition forinhibiting the granulation and/or degranulation of white blood cells,which is intended to be administered topically and externally or byinhalation, such that the first component—the active principle—is anoligosaccharide of formula I containing from three to five saccharideunits and having at least one D-galactose unit linked via an α[1-6] bondto a sucrose unit:

[0018] in which R₁, R₂, R₃ and R₄, independently of each other,represent a hydrogen atom, an alkyl group containing from 1 to 10 carbonatoms and optionally having at least one unsaturation, an acid functioncontaining from 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide, saidoligosaccharide preferably being chosen from the group consisting ofraffinose and stachyose in free or derived form, and

[0019] the second component, which is a molecule that is positivelycharged at physiological pH and that promotes pinocytosis and/or amolecule that promotes membrane penetration.

[0020] Said cosmetic composition may also contain an antiallergiccomponent.

[0021] The preferred combinations for use for the purposes of thepresent invention are the following: the oligosaccharide is raffinose infree or derived form and the second component a saponin; theoligosaccharide is raffinose in free or derived form and the secondcomponent is arginine; the oligosaccharide is stachyose in free orderived form and the second component is a saponin; the oligosaccharideis stachyose in free or derived form and the second component isarginine.

[0022] According to one preferred form, said composition is such thatthe weight concentration of oligosaccharide is between 0.01% and 20%relative to the total mass of the composition and preferably between0.01% and 10% relative to the total mass of the composition. Preferably,the concentration of the second component (saponins and arginine) ispreferably between 5% and 20% by weight relative to the oligosaccharideconcentration and preferably about 10% by weight relative to theoligosaccharide concentration.

[0023] This composition also generally contains at least onecosmetically acceptable excipient.

[0024] The cosmetic compositions used according to the present inventionmay be in the form of a solution, an emulsion, a cream, an ointment, apowder, a milk, a lotion, a gel or a paste with water. Without wishingto be bound by any particular theory, it would appear that ingestion(drinks, lozenges or gel capsules) makes the composition ineffectivesince, quite probably, the enzymatic systems of the stomach destroy thesucrose part of the molecule, converting it into melibiose. However, invitro studies showed that melibiose was not active on leukocytes.

[0025] A subject of the present invention is also a cosmetic treatmentprocess such that said cosmetic composition is applied topically.

[0026] A subject of the present invention is also the use of acombination of at least two components for the manufacture of adermatological composition, i.e. a composition as a medicinal product,to inhibit the granulation and/or degranulation of white blood cells,which is intended to be used externally and topically or by inhalation.This composition is especially intended for treating at least onesymptom possibly associated with allergy, chosen from the groupconsisting of asthma, eczema, pruritus, psoriasis, allergicconjunctivitis, urticaria, reactions to insect bites, and itching, inparticular symptomatic itching of major burn sufferers, or as ananti-inflammatory, in particular for treating arthrosis.

[0027] The present invention relates to the use, for the manufacture ofa medicinal product for inhibiting the granulation and/or degranulationof white blood cells, which is intended to be taken in the form ofointments, creams, eyedrops, aerosols (sprays), lotions or byinhalation, of a composition comprising at least:

[0028] the first component—the active principle—of said combination isan oligosaccharide of formula I containing from three to five saccharideunits and having at least one D-galactose unit linked via an α[1-6] bondto a sucrose unit:

[0029] in which R₁, R₂, R₃ and R₄, independently of each other,represent a hydrogen atom, an alkyl group containing from 1 to 10 carbonatoms and optionally having at least one unsaturation, an acid functioncontaining from 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide, saidoligosaccharide preferably being chosen from the group consisting ofraffinose and stachyose in free or derived form, and

[0030] the second component is a molecule that is positively charged atphysiological pH and that promotes pinocytosis, or a molecule thatpromotes membrane penetration, this second component preferably beingchosen from the group consisting of saponins and arginine.

[0031] The combinations that are preferred for use for the purposes ofthe present invention are as follows: the oligosaccharide is raffinosein free or derived form and the second component is a saponin; theoligosaccharide is raffinose in free or derived form and the secondcomponent is arginine; the oligosaccharide is stachyose in free orderived form and the second component is a saponin; the oligosaccharideis stachyose in free or derived form and the second component isarginine.

[0032] According to one preferred form, said composition is such thatthe weight concentration of oligosaccharide is between 0.01% and 20%relative to the total mass of the composition and preferably between0.01% and 10% relative to the total mass of the composition. Preferably,the concentration of the second component (saponins, arginine) ispreferably between 5% and 20% by weight relative to the oligosaccharideconcentration and preferably about 10% by weight relative to theoligosaccharide concentration.

[0033] Said composition as a medicinal product may also contain anantiallergic component.

[0034] Said medicinal product advantageously contains at least onepharmaceutically acceptable excipient.

[0035] This medicinal product may be intended for local or generaltreatment and may be in the form of a solution, an emulsion, a cream, anointment, a powder, a milk, a lotion, a gel or paste with water,eyedrops or a spray.

[0036] The main indications of this medicinal product are the treatmentsof at least one symptom possibly associated with an allergy chosen fromthe group consisting of asthma, eczema, pruritus, psoriasis, allergicconjunctivitis, urticaria, reactions to insect bites, or itching, inparticular symptomatic itching of major burn sufferers, and thismedicinal product is also recommended as an anti-inflammatory, inparticular for treating arthrosis.

[0037] A subject of the present invention is also a process forpreparing a composition according to the invention such that at leastthe following are mixed together:

[0038] an oligosaccharide of formula I,

[0039] optionally at least one second component, which is a moleculethat is positively charged at physiological pH and that promotespinocytosis, or a molecule that promotes membrane penetration,

[0040] optionally an antiallergic component.

[0041] These oligosaccharides consisting of galactose units linked viaone (or more) bonds of α type between them and to a sucrose unit, arethe storage substances of a certain number of plants (bean, pea,soybean, etc.) and are especially known for their responsibility for thephenomenon of flatulence in man. On the other hand, no direct action ofpossible therapeutic (dermatological) or cosmetic interest has ever beendemonstrated to date.

[0042] Without wishing to be bound by any particular theory, thesemolecules were tested on the antidegranulating activity of humanbasophils, starting from the idea that since mammalian bodies do nothave the enzymes necessary to degrade α-linked galactose units, theseoligosaccharides might disrupt the glycolysis of mastocytes, i.e. theenergy source for granulation and degranulation.

[0043] The oligosaccharides used to carry out the present inventioncomprise from three to five saccharide units, with at least oneD-galactose unit linked via its carbon 1 to carbon 6 of a sucrose unit,via an α bond according to formula I below:

[0044] in which R₁, R₂, R₃ and R₄ represent, independently of eachother, a hydrogen atom, an alkyl group containing from 1 to 10 carbonatoms and optionally having at least one unsaturation, an acid functioncontaining from 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide, which may be infree or derived form.

[0045] In addition, the sucrose unit may also be in free form (that isto say a sugar in which the alcohol functions are unprotected) orderived form. Moreover, this sucrose unit appears to be essential forthe biological activity since melibiose, tested under the sameexperimental conditions, shows absolutely no activity on mastocytes.

[0046] For the purposes of the present invention, the expression“derived form” means that the alcohol functions are substituted, forexample, with an acetyl group (CH₃CO), etc.

[0047] Preferably, stachyose and raffinose will be used.

[0048] Stachyose is a tetraholoside whose final structure is:

[0049]α-D-galactopyranosyl-(1-6)-α-D-galactopyranosyl-(1-6)-α-D-glucopyranosyl-(1-2)-β-D-fructofuranoside.

[0050] Stachyose and raffinose correspond to the formula given below.

[0051] in which, for stachyose, R=H and n=2 and, for raffinose, R=H andn=1.

[0052] The compositions, in particular the cosmetic compositions orcompositions as medicinal (dermatological) product, according to theinvention may be in the form of ointments, creams, eyedrops, sprays orlotions for local administration, in combination with compatibleexcipients. The excipients generally used to prepare such compositionsare binders, preserving agents, flavorings, etc. In these forms, theweight percentage of active principle relative to the mass of the totalcomposition is between 0.01% and 20% and preferably 0.2% and 1%.Pharmaceutical compositions for inhalation may also be prepared forinternal treatments (asthma).

[0053] In order to test the antidegranulating or antiactivating activityof the oligosaccharides on human basophils by flow cytometry, leukocytesuspensions from different individuals selected as being good atresponding to anti-IgE were used to isolate the leukocytes, by simplesedimentation at 1 g. These leukocytes are preincubated for 30 minuteswith successive dilutions from 10 to 0.01 mg/ml of the test product. Thebasophils are then stimulated with a target concentration of a humananti-IgE labeled with a mixture of anti-IgE FITC and anti-CD63PEantibodies, CD63 being an activation label for human basophils. Thepercentages of activation or of inhibition are then calculated by flowcytometry. The biological activity of the oligosaccharides was thusevaluated.

[0054] The chemical structure of the commercial molecules andderivatives, in particular that of stachyose, was confirmed by NMR: ¹Hand ¹³C NMR, 2D-NMR (DQF-cosy, HMQC and HMBC).

[0055] The same biological tests were performed on structural analogssuch as raffinose or oligosaccharides comprising an α bond between agalactose unit and another saccharide unit such as in melibiose.

[0056] These highly water-soluble molecules have little chance ofcrossing the liposoluble membranes of the mastocytes, as confirmed bythe test results.

[0057] However, it is well known that the mastocytes show largepinocytotic activity with respect to molecules that are positivelycharged (i.e. bases) at physiological pH.

[0058] Tests were thus undertaken. The aim of these tests is to test thedegranulation-inhibiting power of compositions containing a firstcomponent—the active principle—which is an oligosaccharide chosen fromstachyose and raffinose at different concentrations and a secondcomponent chosen from the group supplied by arginine, choline, calciumchloride (CaCl₂) and saponin. The percentage of inhibition wascalculated by repeating the experiment three times.

[0059] The results obtained are given in table 1, and the followingabbreviations have been used: “S”: for stachyose “R”: for raffinose“10”: for 10 mg/ml “1”: for 1 mg/ml “0.1”: for 0.1 mg/ml “a”: forarginine “b”: for choline “c”: for CaCl₂ “Sp”: for saponin “NT”: for nottested.

[0060] Composition S₁₀a S₁a S_(0.1)a S₁₀b S₁b S_(0.1)b % of inhibition31 15  0 9 0 0 Composition S₁₀c S₁c S_(0.1)c S₁₀sp S₁sp S_(0.1)sp % ofinhibition  6 0 0 76  0 0 Composition R₁₀a R₁a R_(0.1)a R₁₀b R₁bR_(0.1)b % of inhibition 19 5 0 2 NT NT Composition R₁₀c R₁c R_(0.1)cR₁₀sp R₁sp R_(0.1)sp % of inhibition  0 NT NT 77 10  0

[0061] In the light of these results, it is clearly seen that, at theconcentrations tested, the compositions containing arginine andstachyose or raffinose make it possible to inhibit the degranulation orthe initiation of the granulation system and thus to inhibit the releaseof histamine, bradykinin, serotonin and the chemotactic factors of otherleukocytes.

[0062] In order to obtain a greater inhibition of mastocytes we thenundertook to promote the penetration of these molecules across themembranes by replacing the arginine with saponin (Quillaja bark). Theresults of table 1 show that even higher percentages of inhibition arethen obtained.

[0063] The use of the oligosaccharides according to the presentinvention, in the presence of molecules that are positively charged atphysiological pH (for example such as arginine) and promote pinocytosis,or of molecules that promote membrane penetration (for example saponins)thus make it possible to block or reduce the biological activities ofthe mastocytes, basophils and eosinophils in mammals.

[0064] No cytotoxic activity was observed during the biological studies.Molecules of this type may thus be used beneficially in all thetherapeutic (dermal) and cosmetic fields in which human mastocytes andbasophils (and consequently eosinophils) are involved. Examples that arementioned include pruritus, insect bites, psoriasis, allergicconjunctivitis, urticaria, eczema and asthma.

[0065] The examples that follow do not in any way limit the scope of thepresent invention. Formulation Examples Topical gel Oligosaccharide 0.1g Arginine (or saponins) 0.01 g Methylcellulose 3 g Purified water qs100 g Preserving agents, fragrance qs Fatty cream Oligosaccharide 0.1 gArginine (or saponins) 0.01 g Purified water 5 ml Preserving agents,fragrance qs (applied in unmodified form or as a greasy tulle) EmulsionOligosaccharide 0.2 g Arginine (or saponins) 0.02 g PEG-1500monostearate 5 g PEG-300 monostearate 2 g Fluid liquid paraffin 5 gPurified water qs 100 g Preserving agents, fragrance qs Inhalation Apreparation for inhalation is obtained from the following mixture:Oligosaccharide 100 mg Arginine (or saponins) 10 mg Excipient 10 g

[0066] A standard excipient for an inhalation formulation, for examplesuch as oleic acid, and a propellent gas such as trichlorofluoromethaneor dichlorodifluoromethane, are added to this mixture.

[0067] Greasy cream A (% formulation) Components % Water 54.60 PEG-6 andPEG-32 stearate 12.00 Butyrospermum parkii 10.00 Raffinose 10.00Glycerol 7.00 Liquid paraffin 3.00 Stearic acid 1.20 Saponin 1.00Methylparaben 0.20 Imidazolidinyl urea 0.20 Bisabolol 0.20 Fragrance0.20 Propylparaben 0.10 BHT, BHA, propyl gallate 0.05 and citric acidBenzyl alcohol, methyl- 0.05 chloroisothiazolinone andmethylisothiazolinone

[0068] Antipruriginous Effect on the Recently Epidermized Tegument of aMajor Burn Sufferer

[0069] During the cicatrization phase, major burn sufferers experienceintense itching, even occasionally under heavy antihistamine treatment.

[0070] This study, which lasted one year, performed on 33 patients,demonstrated the powerful antipruriginous effect of composition A inthis indication.

[0071] Protocol

[0072] Patients suffering from itching are treated with an applicationof composition A whose raffinose concentration is 10%, or with anapplication of composition A′ whose raffinose concentration is 5%, oralternatively with an application of composition A″ whose raffinoseconcentration is 3%, the application being attributed at random. Thepercentage of saponin in compositions A′ and A″ represents one tenth ofthe raffinose concentration.

[0073] In the event of a second area to be treated in parallel, thepatient is treated under double blind conditions and without hisknowledge with a composition considered as a placebo on this secondarea, containing the same excipients but not containing any raffinose orsaponin.

[0074] The three tubes with different concentrations of raffinose arecoded for the patient and the staff responsible for applying them.

[0075] The applications are repeated several times a day in the event ofitching and at the request of the patient and may be spread over severaldays.

[0076] A monitoring sheet indicates the time of action of the product(s)according to an analog scale of the pruritus from 0 to 10.

[0077] Results and Conclusions

[0078] Out of 33 cases treated, 30 showed an action of the product, and3 were found to be negative.

[0079] 8 cases involved two areas of application against placebo.

[0080] The average time of action of composition A in the 30 positivecases is between 2 and 5 minutes, with an average duration of action ofbetween 3 and 4 hours.

[0081] The intensity of the initial pruritus estimated on average as 7on the analog scale, is 0 in the 30 cases after 15 minutes.

[0082] No statistically significant difference could be demonstrated asregards the three doses of raffinose.

[0083] Against placebo, the same effects as above are still observed forthe area treated with composition A.

[0084] For the area treated with the placebo, the intensity of thepruritus decreases slightly from 0 to 5 minutes and then returns to theinitial level of intensity after 5 to 15 minutes. This well-knowntransient phenomenon is linked to the moisturization of the skin.

[0085] No intolerance phenomena were reported during the study in singleor repeated application.

[0086] It should be noted that one case of eczema regressed under theeffect of the treatment.

[0087] Out of 3 negative cases, one is unexplained and, as for the othertwo patients, one suffered from a very intense inflammation and theother from a burning sensation, but not really pruritus.

1. The use of a combination of at least two components for themanufacture of a cosmetic or dermatological composition for inhibitingthe granulation and/or degranulation of white blood cells, which isintended to be administered topically and externally or by inhalation,such that the first component, which is the active principle, is anoligosaccharide of formula I containing from three to five saccharideunits and having at least one D-galactose unit linked via an α[1-6] bondwith a sucrose unit:

in which R₁, R₂, R₃ and R₄, independently of each other, represent ahydrogen atom, an alkyl group containing from 1 to 10 carbon atoms andoptionally having at least one unsaturation, an acid function containingfrom 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide, and the secondcomponent is a molecule that is positively charged at physiological pHand that promotes pinocytosis or a molecule that promotes membranepenetration.
 2. The use as claimed in claim 1, such that saidoligosaccharide is chosen from the group consisting of raffinose andstachyose in free or derived form.
 3. The use as claimed in claim 1 or2, such that the second component is chosen from the group consisting ofsaponins and arginine.
 4. The use as claimed in claim 1, such that theoligosaccharide is raffinose in free or derived form and the secondcomponent is a saponin.
 5. The use as claimed in claim 1, such that theoligosaccharide is raffinose in free or derived form and the secondcomponent is arginine.
 6. The use as claimed in claim 1, such that theoligosaccharide is stachyose in free or derived form and the secondcomponent is a saponin.
 7. The use as claimed in claim 1, such that theoligosaccharide is stachyose in free or derived form and the secondcomponent is arginine.
 8. The use as claimed in one of the precedingclaims, characterized in that the weight concentration ofoligosaccharides is between 0.01% and 20% relative to the total mass ofthe composition.
 9. The use as claimed in one of the preceding claims,characterized in that the concentration of the second component isbetween 5% and 20% by weight relative to the oligosaccharideconcentration.
 10. The use as claimed in one of the preceding claims,characterized in that it comprises an antiallergic component.
 11. Theuse as claimed in one of the preceding claims, characterized in that theweight concentration of oligosaccharides is between 0.01% and 10%relative to the total mass of the composition and the concentration ofthe second component is about 10% by weight relative to theoligosaccharide concentration.
 12. The use as claimed in one of thepreceding claims, for the treatment of at least one symptom associatedwith an allergy chosen from the group consisting of asthma, eczema,pruritus, psoriasis, allergic conjunctivitis, urticaria, reactions toinsect bites, and itching, and in particular the symptomatic itching ofmajor burn sufferers, or as an anti-inflammatory, in particular fortreating arthrosis.
 13. The use as claimed in one of the precedingclaims, such that said composition is in the form of an ointment, acream, eyedrops, a lotion, a spray intended for external use, or a sprayfor inhalation.
 14. A cosmetic treatment process for inhibiting thegranulation and/or degranulation of white blood cells, characterized inthat a composition is applied topically, this composition comprising atleast one first component, which is the active principle, which is anoligosaccharide of formula I containing from three to five saccharideunits and having at least one D-galactose unit linked via an α[1-6] bondto a sucrose unit:

in which R₁, R₂, R₃ and R₄, independently of each other, represent ahydrogen atom, an alkyl group containing from 1 to 10 carbon atoms andoptionally having at least one unsaturation, an acid function containingfrom 1 to 10 carbon atoms and optionally having at least oneunsaturation, a sulfate function or a monosaccharide, and at least onesecond component, which is a molecule that is positively charged atphysiological pH and that promotes pinocytosis, or a molecule thatpromotes membrane penetration and at least one pharmaceuticallyacceptable excipient.